We have developed a powerful molecular system for detection and identification of bacterial strains (clonal types). This system makes it possible to answer important questions about the periodontitis-associated bacteria Actinobacillus actinomycetemcomitans (Aa) and Porphyromonas gingivalis (Pg) that could not be addressed until efficient, precise strain identification technology was developed. A polymerase chain reaction (PCR)-based molecular analysis using species-specific primers will be used to identify strains of both microbes. It is not necessary to isolate and culture the bacteria with this approach. To determine the genetic identity for individual clonal types, we will amplify by PCR and sequence the highly variable transcribed spacer region between the 16S and 23S ribosomal genes directly from plaque. To determine clonal type matches for transmission studies we will use mutation detection enhancement (MDE) gel electrophoresis analysis of the spacer region fragment. Little is known about the genetic diversity of strains of Aa and Pg that might be found within human populations. We will investigate the level of recombination between strains to determine if one or only a few markers are sufficient for genetic studies. We will determine the genetic population structure of Aa and Pg at several locations in North America and around the world. Research on strains of Aa and Pg is conducted at many geographic locations, and there is currently no universal system for identification of strains. Our approach will allow us to catalog the major strains and can provide a link between studies performed by different research groups. We have proposed studies of extended family groups to allow comparisons of transmission rates to be made between groups with different degrees of intimacy and longevity of contact. These comparisons will allow inferences about the ease of transmission of bacteria between contacts and the long-term stability of infection. More information about transmission patterns between intimate associates is important for planning prevention and treatment strategies for what may be potentially contagious bacterial infections, and for planning future studies to examine the mechanisms and factors involved in the transmission of these bacteria. We have also proposed studies in which the clonal type distribution of Aa and Pg in a population with indicators of disease will be compared to the clonal type distribution found in a healthy control population. If we find that the population of strains colonizing the diseased group is different than that found in healthy individuals, this would indicate that these disease-associated strains are pathogens in the classic sense. If distributions are found to be the same in the 2 groups, this would imply that the bacteria are opportunistic pathogens. The discovery of pathogenic strains would have immediate implications for devising diagnosis, treatment and prevention strategies and would facilitate further molecular genetic investigation into the pathogenic process.